Stem cells are undifferentiated cells that exist in many tissues of embryos and adult mammals. Both adult and embryonic stem cells are able to differentiate into a variety of cell types and, accordingly, may be a source of replacement cells and tissues that are damaged in the course of disease, infection, or because of congenital abnormalities. (See, e.g., Lovell-Badge Nature 2001, 414:88-91; Donovan et al. Nature 2001, 414:92-97). Various types of putative stem cells exist which; when they differentiate into mature cells, carry out the unique functions of particular tissues, such as the heart, the liver, or the brain. Pluripotent stem cells are thought to have the potential to differentiate into almost any cell type, while multipotent stem cells are believed to have the potential to differentiate into many cell types (Robertson, Meth. Cell Biol. 75:173,1997; and Pedersen, Reprod. Fertil. Dev. 6:543, 1994).
However, certain cell types (such as nerve cells and cardiac cells) differentiate during development and adult organisms do not replace these cells. It would be of particularly great value in treating a wide variety of diseases to have renewable sources of stem cells that can reliably differentiate into the desired phenotype. By way of example, Parkinson's Disease (PD) is a progressive degenerative disorder that appears to be associated with the death of dopamergic neurons extending from the substantia nigra of the brain into the neighboring striatum. Attempts to treat PD by transplanting stem cells collected from the developing brains of aborted fetuses have had mixed results. (See, e.g,. Freed et al. (2001) N. Engl. J. Med. 344:710-719). Further, ethical considerations have mitigated against the use of these embryonic or fetal stem cells. Additionally, it has proven difficult to discover conditions under which embryonic or adult stem cells differentiate into the desired phenotype.
Furthermore, even in those cell types, such as epithelial cells and hematopoietic cells, that are replaced in adult organisms it has been a significant challenge to readily and inexpensively obtain stem cells in significant quantities. For example, mammalian hematopoietic cells (e.g., lymphoid, myeloid and erythroid cells) are all believed to be generated by a single cell type called the hematopoietic “stem cell.” (Civin et al. (1984) J. Immunol. 133:157-165). However, these hematopoietic stem cells are very rare in adults, accounting for approximately 0.01% of bone marrow cells and isolation based on cell surface proteins such as CD34 results in very small yields. Schemes to fractionate human hematopoietic cells into lineage committed and non-committed progenitors are technically complicated and often do not permit the recovery of sufficient cells to address multilineage differentiation. (see, e.g., Berenson et al., 1991; Terstappen et al., 1991; Brandt et al. (1988) J. Clinical Investigation 82:1017-1027; Landsdorp and Dragowska (1992) J. Exp. Med. 175:1501-1509; Baum et al. (1992) Proc. Natl. Acad. Sci. 89:2804-2808).
Similarly, existing protocols that induce differentiation ex vivo exert little control over cell fate, thereby yielding diverse and impure cell populations that are inadequate for projects involving ex vivo reconstitution of the immune system. (See, e.g., Clarke et al. Science 2000, 288:1660-1663; Bjornson et al. Science 1999, 283:534-537; Galli et al. Nat Neurosci 2000, 3:986-991; Mezey et al. Science 2000, 290:1779-1782; Toma et al. Nat Cell Biol 2001, 3:778-784; Weissman et al. Annu Rev Cell Dev Biol 2001, 17:387-403; Anderson et al. Nat Med 2001, 7:393-395; Morrison Curr Biol 2001, 11:R7-9; Lagasse et al. Nat Med 2000, 6:1229-1234; Krause et al. Cell 2001, 105:369-377). In addition, certain existing protocols for stem cell growth and differentiation are dependent on the use of feeder cells which necessitates the efficient scale-up of cell culture and creates associated risks including, infection, cell fusion and/or contamination.
Therefore, although embryonic stem cells (ES cells) can be maintained in culture in an undifferentiated state, ex vivo conversion to a desired cell type is difficult. See, e.g., Clarke et al. Science (2000) 288:1660-1663. Similarly, adult stem cells are very difficult to expand in culture. See, e.g., Reya et al. Nature 2001, 414:105-111; Tang et al. Science 2001, 291:868-871.
Thus, there is a clear need to develop methods for identifying, propagating and altering the state (e.g., by differentiation or dedifferentiation) of stem cells to provide a source of cells that are transplantable to the CNS, PNS, or other tissues in vivo in order to replace damaged or diseased tissue.